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Notes: Abstract The DNA repair host-mediated assay, in which repairable DNA damage is determined in E. coli cells present in various organs of mice exposed to genotoxic agents, was further developed to broaden the range of organs under study and to simplify the procedure of assessing differential bacterial cell survival. A pair of derivatives of E. coli K-12 strain 343/113 was constructed which differed vastly in DNA repair capacity (uvr +/rec + vs uvrB/recA), as a means of assessing DNA damaging effects; furthermore, the strains differed in their ability to ferment lactose (Δ Lac vs Lac +), so that the individual survival of both strains could be determined on a single agar medium (containing neutral red as pH indicator), on which the strains had different colony colour morphology (red, Lac + vs white, Lac − colonies). Finally, the strains were made streptomycin-dependent, to prevent uncontrolled growth of the bacterial cells within the various organs and also to inhibit contamination of the survival agar medium by representatives of the normal intestinal microflora. The experimental procedure consisted of injecting mixtures of stationary cells of the two strains (ca. 3–5×108 viable cells per mouse) both intravenously and orally into mice, either pretreated or subsequently treated with test chemicals. Ninety minutes after injection of the bacteria, the liver, spleen, lungs, kidneys, stomach, intestine, colon, and ca. 50 μl blood, were removed, suspended in buffer, homogenized, and the survival of the two strains determined on neutral red agar supplemented with streptomycin. In preliminary experiments in which the mice were treated with intraperitoneal injections of mitomycin C (0–2.0 mg per kg body weight), a dose-dependent increase in DNA damaging activity was induced in bacterial cells present in all organs tested, the lowest effects being observed in kidneys and lungs, and the highest in liver and blood. These results need further confirmation in more extensive tests, but they do nevertheless clearly indicate the possible usefulness of the DNA repair host-mediated assay as a rapid biological dose monitor for obtaining information on the genotoxic activity in vivo of compounds for which long-term mutagenicity and carcinogenicity data are not yet available.

Notes: Abstract The introduction of foreign cells (e.g. ascites tumor cells) into laboratory mammals and their subsequent recovery after treatment of the host with exogenous chemicals to determine the induction of genetic effects (e.g. chromosomal aberrations) is a technique which has been employed for more than 20 years. The use of bacteria as indicators of induced point mutations was first described by Legator et al. (1969). In their technique which they called a host-mediated assay, the microbes (e.g. Salmonella typhimurium) were injected into the peritoneal cavity of mice and, thus, exposed to potentially mutagenic metabolites of the compound under test. Today a wide variety of genetical changes can be detected in several indicator organisms. In addition to the histidine-requiring strains of Salmonella which allow the detection of different types of back-mutations, some other enterobacteria have proven useful. Auxotrophic strains of Serratia marcescens to detect back mutations, and Escherichia coli bacteria in which both forward and back mutations can be assayed simultaneously in several different genes. With fungi or fungal spores as indicators further effects of genetical importance can be determined, e.g. the consequences of recombination processes such as mitotic gene conversion and mitotic recombination in the yeasts, and of deletions in Neurospora crassa (conidia). After the successful development of methods to measure the induction of point mutations in cultured mammalian cells, it is also possible now to use established animal cell lines (e.g. mouse lymphoma cells) in a host-mediated assay provided isogenic or compatible hosts are available. During the past several years the route of administration of the indicator cells and the distribution of these cells within the animal body have been studied. Efforts have been made to bring the indicators into close contact with the reproductive organs or within the liver because this is the organ where most foreign compound metabolism is known to occur. Depending on the inoculation technique (in situ, intraperitoneal, intravenous) it is now possible to recover indicators out of testes, liver spleen, lungs and peritoneal cavity of treated animals in quantities large enough to perform genetic tests. A further improvement is the introduction of indicators within the intestinal tract of rodents; the first experiments along this line using Salmonella seem promising and have opened the way toward using common representatives of the intestinal flora such as Escherichia coli. In general it appears that the host-mediated assay technique is a useful tool to use in assessing the degree of mutagenicity and the organospecificity of foreign chemicals in living mammals and that it should be used until techniques that allow the detection of genetic events in the cells of the different organs have been developed. In the near future, host-mediated assays will have to be used to assess more quantitatively the mutagenicity of the numerous chemicals that have been found genetically active by using direct microsomal assays. Further improvements of host-mediated assays are necessary. For instance, there is as yet no practical method for retention of microbial indicators long enough within the blood stream of living animals to perform genetic tests. This would be desirable for pharmaceutical reasons, e.g. for determination of the kinetics of appearance and removal of mutagenic factors within the blood stream. Preliminary experiments indicate that bacteriophages might be useful indicators, since they are present in blood in quantities sufficient for genetic analysis more than 24 h after intravenous injection. In contrast to all other indicators used so far, bacteriophages do not show any metabolism outside their bacterial hosts that might interfere with the mammalian metabolism. Studies are presently under way to determine what types of mutations or of genetic alterations are best detected after extracellular phage treatment.

Notes: Objective To compare the tocolytic potency of ritodrine, nicardipine and atosiban, used alone and in dual combinations, to see whether combinations of these drugs, which act via different pathways, could improve inhibition of uterine contractility.Design Study on myometrial contractility in vitro.Setting Laboratory of physiology, Lyon, France.Sample Longitudinal myometrial strips from non-labouring timed pregnant Wistar rats (18 gestational days).Methods Strips were simultaneously exposed to EC25, EC50 or EC75 of dual combinations of either ritodrine and nicardipine, ritodrine and atosiban or nicardipine and atosiban (n= 10/group). Basal contractile activity and contractile activity after addition of each combination was measured using the 10 min integral of activity. Changes were expressed as percentage from the basal 10 min integral activity. The observed percentage inhibition of activity was compared with the expected percentage inhibition in an additive pharmacological model. When no significant difference occurred, the combination was deemed simply to have an additive tocolytic effect. When inhibition of activity was significantly greater compared with the expected percentage inhibition, the combination was deemed to have a synergistic effect.Main outcome measure Changes in integral contractile activity in response to tocolytic combinations.Results Ritodrine and atosiban inhibited integral activity to a greater extent than expected [e.g. using EC50: observed inhibition 88.9% (13.8%) vs expected inhibition 75%; P 〈 0.015]. Actual inhibition by nicardipine/ritodrine [78.55% (20.4%) vs 75%; P= n.s.] and nicardipine/atosiban [78.94% (17.8%) vs 75%; P= n.s.] was not significantly different from expected.Conclusions A combination of ritodrine plus atosiban exhibits a synergistic inhibition for myometrial activity, thus allowing the use of lower concentrations of each drug to achieve the same effect compared with each drug used alone. Combination of nicardipine plus ritodrine and nicardipine plus atosiban achieves only an additive effect. The potential for decreasing side effects (beta-mimetics) and costs (atosiban) when using a combination in clinical practice needs to be evaluated.